Biology subjects

Lopez, B.

Publications and source records attributed to Lopez, B..

10 recordsLinked to original sources

bioRxiv2026

Integrated Skin Sensitization Assessment Using Consensus Modelling and Tiered New Approach Methodology: A SaferSkin Case Study

The transition toward animal-free safety assessment of chemicals has accelerated the development of New Approach Methodologies (NAMs) for predicting skin sensitization. However, individual in silico models and experimental NAM assays frequently produce inconsistent or contradictory results, limiting their reliability when used in isolation. To address this challenge, we present a tiered integrated assessment framework implemented through the open source SaferSkin application, which enables systematic comparison and integration of multiple predictive models and experimental data within a transparent weight-of-evidence workflow. In this case study, a diverse set of 21 reference compounds was evaluated using a battery of in silico approaches, including the OECD QSAR Toolbox, VEGA, CASE Ultra and additional machine-learning models implemented within SaferSkin. The platform enables side-by-side comparison of predictions and integration of experimental data through Bayesian network models, allowing probabilistic updating of predictions as new evidence becomes available. Our results demonstrate that reliance on any single predictive model is insufficient for robust hazard identification due to frequent disagreement between models. In contrast, consensus interpretation across multiple modelling approaches combined with targeted experimental evidence substantially improves predictive confidence. The integrated weight-of-evidence framework showed strong concordance with reference classifications and was further supported by independent validation using the Pred-Skin Bayesian model. Importantly, the tiered workflow enables resolution of ambiguous cases. For example, lower-tier predictions for ethyl (2E,4Z)-deca-dienoate were inconsistent across models, whereas targeted third-tier testing using the SENS-IS assay identified the compound as a strong sensitiser (GHS Category 1A). Overall, this study demonstrates how integrated modelling, Bayesian evidence updating and targeted NAM testing can reduce uncertainty in skin sensitization assessment. The SaferSkin framework provides a transparent and reproducible approach for implementing Next Generation Risk Assessment (NGRA) strategies and supports the development of animal-free regulatory toxicology and Safe-and-Sustainable-by-Design chemical innovation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/711911v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@196a52eorg.highwire.dtl.DTLVardef@13690a6org.highwire.dtl.DTLVardef@1528703org.highwire.dtl.DTLVardef@11e05d2_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO C_FIG

pharmacology and toxicology↗

bioRxiv2025

Anti-vitronectin single-chain variable fragment targeting the hemopexin-like domain decreases age-related macular degeneration indicators in ex vivo models of the retinal pigment epithelium

Vitronectin is a prominent constituent of the extracelluar matrix (ECM) that plays a key role in inflammation by regulating cell adhesion, migration, and complement activation. Together with other inflammatory biomarkers, lipids, and hydroxyapatite, an ECM component, vitronectin, accumulates in abnormal deposits and drusen associated with age-related macular degeneration (AMD). Using an engineered polypeptide (VnHX) containing the hexmopexin-like domain (HX) of vitronectin, we found that the HX domain directly promotes hydroxyapatite (HAP) accumulation. Next, we screened humanized single-chain variable fragments (scFvs) against vitronectin that inhibit the interaction between VnHX and HAP, leveraging the binding of VnHX to HAP. We then tested these scFvs for their ability to block Vn-containg deposits secreted by stem cell-derived retinal pigment epithelium (RPE). In this in vitro drusen model, treatment with the anti-vitronectin antibodies decreased vitronectin accumulation. Furthermore, the antibody treatment led to decreased accumulation of C5b-9 and clusterin, indicating alterations in the complement pathway and cellular stress. These results support that vitronectin has a functional role drusen accumulation and possibly in AMD progression. Vitronectin would be a novel, promising therapeutic target for AMD.

cell biology↗

bioRxiv2024

Skin Sensitisation Case Study: Comparison of Defined Approaches including OECD 497 Guidance

Characterising known and new chemical compounds for skin sensitisation provides a basis for the development of safer products where ingredients are exposed to skin. By including new approaches, such as tiered testing strategies and integrated data analysis, it is possible to develop next generation products adhering to emerging regulations, scientific evidence and animal welfare principles. To ensure data integrity during such assessment the OECD provides characterisation guidelines and Defined Approaches (DAs) to uniform work-flows. In this study we developed and applied the integrated characterisation tool called < > to compare the results of different DAs for eight compounds and included results obtained from current OECD guidance and emerging methods. We tested two compounds with unclear or indeterminate results with the SENS-IS assay to explore the value of the experiment in strengthening the weight of evidence and arriving at a clearer conclusion.

pharmacology and toxicology↗

bioRxiv2022

Dendritic cell-Natural Killer cell Crosstalk Modulates T cell activation in Response to Influenza A Viral Infection

Influenza viruses lead to substantial morbidity and mortality including ~3-5 million cases of severe illness and ~290,000-650,000 deaths annually. One of the major hurdles regarding influenza vaccine efficacy is generating a durable, robust cellular immune response. Appropriate stimulation of the innate immune system is key to generating cellular immunity. Crosstalk between innate dendritic cells (DC) and natural killer (NK) cells plays a key role in activating virus-specific T cells, yet the mechanisms used by influenza A viruses (IAV) to govern this process remain incompletely understood. Here, we used an ex vivo autologous human primary immune cell culture system to evaluate the impact of genetically distinct IAV strains on DC-NK cell crosstalk and subsequent T cell activation. We report that the addition of NK cells to cultures containing both DCs and naive T cells led to an increase in the frequency of CD69+ and CD25+ T cells and elevated levels of IFN-{gamma}, TNF, and IL-10. However, upon IAV infection of DCs, the addition of NK cells to cultures no longer increased the frequency of CD25+ T cells nor elevated IFN-{gamma}, TNF, and IL-10 cytokine levels. Investigation of the impact of IAV infection on DC-NK crosstalk revealed that exposure of DCs to influenza virus in co-culture led to an increased frequency of HLA-DR+ and a decreased frequency of CD83+ and CD86+ cells-molecules involved in stimulating T cell activation. An expansion of an HLA-DR+ NK cell subset was observed following culture with influenza-infected DCs in a contact-dependent and cytokine independent-manner. Overall, our results indicate a role for DC-NK cell crosstalk in T cell priming in the context of influenza infection, informing the immunological mechanisms that could be manipulated for the next generation influenza vaccine or immunotherapeutic.

immunology↗

bioRxiv2022

Human RAD52 stimulates the RAD51-mediated homology search

Homologous recombination (HR) is a DNA repair mechanism of double strand breaks and blocked replication forks that involves a process of homology search and homologous pairing leading to the formation of synaptic intermediates whose architecture and dynamics are tightly regulated to ensure genome integrity. In this mechanism, RAD51 recombinase plays a central role and is supported by many partner including BRCA2 and RAD52. If the mediator function of BRCA2 to load RAD51 on RPA-covered ssDNA is well established, the role of RAD52 in HR, more precisely its interplay with BRCA2 is still far from understood. We have used Transmission Electron Microscopy combined with biochemistry to characterize the sequential participation of RPA, RAD52 and BRCA2 in the assembly of the RAD51 filament, its architecture and its activity. Despite our results confirm that RAD52 lacks a mediator activity, we observed that RAD52 can tightly bind to RPA-coated ssDNA, inhibit the mediator role of BRCA2 and form shorter RAD52- and RAD51-containing mixed filaments that are more efficient in subsequent homology search and formation of synaptic complexes and D-loops, resulting in more frequent multi-invasions as well. Through the characterization of the behavior of RAD52 and BRCA2, these results provide new molecular insights on the formation and regulation of presynaptic and synaptic intermediates during human HR.

biochemistry↗

bioRxiv2021

Receptor-binding domain recombinant protein RBD219-N1C1 on alum-CpG induces broad protection against SARS-CoV-2 variants of concern

We conducted preclinical studies in mice using a yeast-produced SARS-CoV-2 RBD subunit vaccine candidate formulated with aluminum hydroxide (alum) and CpG deoxynucleotides. This formulation is equivalent to the Corbevax vaccine that recently received emergency use authorization by the Drugs Controller General of India. We compared the immune response of mice vaccinated with RBD/alum to mice vaccinated with RBD/alum+CpG. We also evaluated mice immunized with RBD/alum+CpG and boosted with RBD/alum. Mice were immunized twice intramuscularly at a 21-day interval. Compared to two doses of the /alum formulation, the RBD/alum+CpG vaccine induced a stronger and more balanced Th1/Th2 cellular immune response, with high levels of neutralizing antibodies against the original Wuhan isolate of SARS-CoV-2 as well as the B.1.1.7 (Alpha), B. 1.351 (Beta), B. 1.617.2 and (Delta) variants. Neutralizing antibody titers against the B.1.1.529 (BA.1, Omicron) variant exceeded those in human convalescent plasma after Wuhan infection but were lower than against the other variants. Interestingly, the second dose did not benefit from the addition of CpG, possibly allowing dose-sparing of the adjuvant in the future. The data reported here reinforces that the RBD/alum+CpG vaccine formulation is suitable for inducing broadly neutralizing antibodies against SARS-CoV-2 including variants of concern.

molecular biology↗

bioRxiv2020

Plasma mass spectrometry fingerprints induced by bacterial endotoxins in murine models as markers of pathophysiological evolution in sepsis

Sepsis has been called "the graveyard of pharmaceutical companies" due to the numerous failed clinical trials. The lack of tools to monitor the immunological status in sepsis constrains the development of immunomodulatory therapies. Here, we evaluated a test based on whole plasma peptidome acquired in a MALDI-TOF-mass spectrometer used for bacterial biotyping and machine-learning algorithms to discriminate the different immunological phases of sepsis. In this proof of concept study, two discrete lipopolysaccharide-(LPS) induced murine models emulating the pro- and anti-inflammatory phases that occur during sepsis were evaluated. The LPS group was inoculated with a single high dose of LPS, recalling the proinflammatory phase, and the IS groups was subjected to increasing doses of LPS to induce the anti-inflammatory/immunosuppression phase. Unstimulated mice served as controls. Both experimental groups showed the hallmarks of pro- and anti- inflammatory phases respectively; the LPS group showed leukopenia and higher levels of cytokines and tissue damage markers, and the IS group showed neutrophilia, lymphopenia and significantly lower antibody titers upon immunization. Principal component analysis of the plasma peptidomes formed three discrete clusters that mostly coincided with the experimental groups. In addition, machine-learning algorithms discriminated the different experimental groups with a sensitivity and specificity of up to 95.7% and 90.9% respectively. Our data reveal the potential of plasma peptidome analysis by MALDI-TOF-mass spectrometry as a simple, speedy and readily transferrable method for sepsis patient stratification that would contribute to therapeutic decision-making based on their immunological status.

immunology↗

bioRxiv2019

Mycobacterium tuberculosis Modulates the Metabolism of Alternatively Activated Macrophages to Promote Foam Cell Formation and Intracellular Survival

The ability of Mycobacterium tuberculosis (Mtb) to persist inside host cells relies on metabolic adaptation, like the accumulation of lipid bodies (LBs) in the so-called foamy macrophages (FM). Indeed, FM are favorable to Mtb. The activation state of macrophages is tightly associated to different metabolic pathways, such as lipid metabolism, but whether differentiation towards FM differs between the macrophage activation profiles remains unclear. Here, we aimed to elucidate if distinct macrophage activation states exposed to a tuberculosis-associated microenvironment can accumulate LBs, and its impact on the control of infection. We showed that signal transducer and activator of transcription 6 (STAT6) activation in interleukin (IL)-4-activated human macrophages (M(IL-4)) prevents FM formation induced by pleural effusion from patients with tuberculosis. In these cells, LBs are disrupted by lipolysis, and the released fatty acids enter the {beta}-oxidation (FAO) pathway fueling the generation of ATP in mitochondria. We demonstrated that inhibition of the lipolytic activity or of the FAO drives M(IL-4) macrophages into FM. Also, exhibiting a predominant FAO metabolism, mouse alveolar macrophages are less prone to become FM compared to bone marrow derived-macrophages. Upon Mtb infection, M(IL-4) macrophages are metabolically re-programmed towards the aerobic glycolytic pathway and evolve towards a foamy phenotype, which could be prevented by FAO activation or inhibition of the hypoxia-inducible factor 1-alpha (HIF-1)-induced glycolytic pathway. In conclusion, our results demonstrate a role for STAT6-driven FAO in preventing FM differentiation, and reveal an extraordinary capacity by Mtb to rewire metabolic pathways in human macrophages and induce the favorable FM. IMPORTANCETuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (Mtb). While its treatment was already standardized, TB remains one of the top 10 death causes worldwide. A major problem is the efficient adaptation of Mtb to the macrophage intracellular milieu, which includes deregulation of the lipid metabolism leading to the formation of foamy macrophages (FM) which are favorable to Mtb. A critical aspect of our work is the use of tuberculous pleural effusions (TB-PE) -- human-derived biological fluid capable of mimicking the complex microenvironment of the lung cavity upon Mtb infection -- to study the FM metabolic modulation. We revealed how the STAT6 transcription factor prevents FM formation induced by PE-TB, and how Mtb counteracts it by activating another transcription factor, HIF-1, to re-establish FM. This study provides key insights in host lipid metabolism, macrophage biology and pathogen subversion strategies, to be exploited for prevention and therapeutic purposes in infectious diseases.

immunology↗

bioRxiv2018

Engineered orthogonal quorum sensing systems for synthetic gene regulation

Gene regulators that are controlled by membrane-permeable compounds called Homoserine lactones (HSLs) have become popular tools for building synthetic gene networks that coordinate behaviors across populations of engineered bacteria. Synthetic HSL-signaling systems are derived from natural DNA and protein elements from microbial quorum signaling pathways. Crosstalk, where a single HSL can activate multiple regulators, can lead to faults in networks composed of parallel signaling pathways. Here, we report an investigation of quorum sensing components to identify synthetic pathways that exhibit little to no crosstalk in liquid and solid cultures. In previous work, we characterized the response of a single regulator (LuxR) to ten distinct HSL-synthase enzymes. Our current study determined the responses of five different regulators (LuxR, LasR, TraR, BjaR, and AubR) to the same set of synthases. We identified two sets of orthogonal synthase-regulator pairs (BjaI/BjaR + EsaI/TraR and LasI/LasR + EsaI/TraR) that show little to no crosstalk when they are expressed in Escherichia coli BL21. These results expand the toolbox of characterized components for engineering microbial communities.

synthetic biology↗

bioRxiv2018

Frequent Monitoring of C-peptide Levels in Newly Diagnosed Type 1 Subjects Using Dried Blood Spots Collected at Home

ObjectiveTo evaluate a novel approach to measure {beta}-cell function by frequent testing of C-peptide concentrations in dried blood spots (DBS)\n\nPatientsThirty-two children, aged 7-17 years, recently diagnosed with type 1 diabetes.\n\nDesignMixed-meal-tolerance-test (MMTT) within 6 and again 12 months after diagnosis with paired venous and DBS C-peptide sampling at 0 and 90 minutes. Weekly DBS C-peptide before and after standardized breakfasts collected at home.\n\nResultsDBS and plasma C-peptide levels (n=115) correlated strongly (r=0{middle dot}91; p<0.001). The Bland-Altman plot indicated good agreement. The median number of home-collected DBS cards per participant was 24 over a median of 6.9 months. Repeated DBS C-peptide levels varied considerably within and between subjects. Adjustment for corresponding home glucose measurements reduced the variance permitting accurate description of changes over time. The correlation of the C-peptide slope over time assessed by repeated home DBS versus area under the curve during the two MMTTs was r=0{middle dot}73; p<0.001. Mixed models showed that a 1-month increase of diabetes duration was associated with 17 pmol/l decline in fasting DBS C-peptide, whereas increases of 1 mmol/l in glucose, 1 year older age-at-diagnosis and 100 pmol/l higher baseline plasma C-peptide were associated with 18, 17 and 61 pmol/l higher fasting DBS C-peptide levels, respectively. In addition, glucose responsiveness decreased with longer diabetes duration.\n\nConclusionOur approach permitted frequent assessment of C-peptide, making it feasible to monitor {beta}-cell function at home. Evaluation of changes in the slope of C-peptide using this method may permit short-term evaluation of promising interventions.

biochemistry↗